DNA-binding activity of the A-factor receptor protein and its recognition DNA sequences

The A-factor receptor protein (ArpA) containing an alpha-helix-turn-alpha-helix DNA-binding consensus sequence at its N-terminal portion plays a key role in the regulation of secondary metabolism and cell differentiation in Streptomyces griseus. A binding site forming a palindrome 24bp in length was initially recovered from a pool of random-sequence oligonucleotides by rounds of a binding/immunoprecipitation/amplification procedure with histidine-tagged ArpA and anti-ArpA antibody. By means of further binding/gel retardation/amplification experiments on the basis of the recovered sequence, a 22 bp palindromic binding site with the sequence 5'-GG(T/C)CGGT(A/T)(T/C)G(T/G)-3' as one half of the palindrome was deduced as a consensus sequence recognized and bound by ArpA. ArpA did not bind to the binding site in the presence of its ligand, A-factor. In addition, exogenous addition of A-factor to the ArpA-DNA complex induced immediate release of ArpA from the DNA. All of these data are consistent with the idea, obtained from previous genetic studies, that ArpA acts as a repressor-type regulator for secondary metabolism and cellular differentiation by preventing the expression of a certain key gene(s) during the early growth phase. A-factor, produced in a growth-dependent manner, releases ArpA from the DNA, thus switching on the expression of the key gene(s), leading to the onset of secondary metabolism and aerial mycelium formation.